Verification device for optical clinical assay systems

ABSTRACT

A device and method for verifying correct performance of an optical clinical assay system is provided.

[0001] This application is a continuation of, and claims the benefit ofpriority from U.S. application Ser. No. 09/138,824, filed Aug. 24, 1998and (provisional) application No. 60/057,903, filed on Sep. 4, 1997, thefull disclosures of which are incorporated herein by reference.

BACKGROUND OF THE INVENTION

[0002] Increasingly in modern clinical chemistry, whole blood samples,often obtained by finger stick methods, are analyzed using automatedautomatic analysis systems (meters) which employ disposable (oftenone-time use) test elements, and a non-disposable electronic test devicethat analyzes the reaction occurring in the whole blood sample in thedisposable test element, and then outputs an answer. Such systems areused for analyzing whole blood samples for glucose, cholesterol, andincreasingly, more complex tests such as coagulation testing(prothrombin time, activated partial thromboplastin time), enzymaticanalytes, and the like.

[0003] Because the answer from these devices are often used to make aclinical decision that can significantly impact the health andwell-being of a patient, verification methods to insure that theanalytical devices are performing correctly are of obvious importance.

[0004] One common method for verifying correct performance of a clinicalanalytical system is through the use of control solution, which isusually a liquid chemical solution with known reactivity. If theanalytical device gives the correct answer with a known referencechemical, then the overall performance of the system can be assessed.

[0005] With modern one-shot, disposable test elements, however, theproblem with liquid control testing is that it is destructive. Thedisposable test element has been destroyed as a result of the testing,and only the now-validated meter now survives to test the actual sample.For this reason, modern verification methods tend to shift validation ofdisposable test elements to manufacturers, who validate batches ofdisposable test elements by statistical sampling methods. The problem ofmeter verification remains, however. Meters are typically used foryears, and can be exposed to environmental extremes, misuse, andmechanical shock.

[0006] Because meter verification using liquid control devices anddisposable test units is an expensive process, and because test unitverification is inherently best suited to statistical lot testing by themanufacturer, there is a need for low-cost methods that can verify theperformance of the is meter without the use of liquid control solutionand disposable test units.

[0007] Analytical devices for temperature sensitive enzymatic analytes,such as blood coagulation, typically have a temperature controlledreaction stage, means to determine the start of the enzymatic reaction,optical means to access the progress of the reaction, and computationalmeans (typically a microprocessor or microcontroller) to interpret theprogress of the reaction and output an answer. To completely verify theperformance of the analysis system, each subsystem must be assessed. Thetemperature controlled reaction stage must be tested for propertemperature control, the means to determine the start of the enzymaticreaction must be tested for proper sensitivity, the optical means toaccess reaction progress must be tested (light source, light detector,integrity of optical stage, etc.), and finally the computational meansmust be tested. Alternatively partial verification of some of thesubsystems may be done, and the remainder of the subsystems tested byalternate means, such as liquid control solution and a disposable testunit.

[0008] To verify the function of such analytical devices, electronicverification or “control” devices or circuits are commonly used. Suchverification devices can simulate the action of an enzymatic sampleinteracting with a disposable reagent. If the analytical device returnsthe proper answer after analysis of the verification device, then theproper functioning of the analytical device can be verified without theexpense of using the one time use reagent cartridges.

[0009] The use of reference paint chips to calibrate and verifyphotometric devices has long been known in the art. When applied to homeblood glucose monitors, such reference chips are often referred to as“check strips”. For example, the LifeScan One-Touch™ blood glucosemonitor includes a calorimetric “check strip” in with its meter system.This “check strip” consists of an opaque plastic strip with a paint chipof known calorimetric properties affixed to it. The check strip isinserted into the meter, and is used to verify the performance of themeter's colorimetric photodetector. The system does not vary theintensity of the calorimetric paint chip target as a function of time tosimulate a normal test reaction, nor does it incorporate means tomonitor the analytical devices' temperature.

[0010] Recent refinements to the basic “paint chip” technique, suitablefor clinical reagents and instrumentation, include U.S. Pat. Nos.4,509,959; 4,523,852; 4,729,657; 5,151,755; 5,284,770; and 5,592,290.U.S. Pat. No. 4,509,959 disclosed an apparatus incorporating many suchreference color chips. U.S. Pat. No. 4,523,852 disclosed a referencestandard, suitable for visually read diagnostic reagent test strips,consisting of many colored reference areas of differing hues. U.S. Pat.No. 4,729,657 disclosed photometer calibration methods using two or morereflectance standards and using least squares regression line analysisto construct and store calibration curves in the analytical device'smemory. U.S. Pat. No. 5,151,755 disclosed methods to detect defects inbiochemical analysis apparatuses measurement means by irradiating areference density plate with light that has passed through a pluralityof interference filters and comparing the relative amounts of reflectedlight obtained by these different measurements. U.S. Pat. No. 5,284,770disclosed use of a check strip, along with an analytical instrumenthaving a user insertable key (memory chip) containing the parameters ofacceptable check strip performance, so that correct instrumentperformance can be automatically verified. U.S. Pat. No. 5,592,290disclosed optical analyzer instrument error correction methods usingstandard color plates incorporating dyes with absorption spectrumsimilar to the analytical reagent normally read by the analyzer. Thesestandard color plates are then used in conjunction with a secondreference optical analyzer and a specific correction algorithm tocorrect the instrument error in the first instrument.

[0011] In addition to passive “paint chip” verification methods, anumber of different active (typically electronic) verification methodshave also been used. These active verification methods typically involveelectronic components, and often produce a dynamic (as opposed to astatic) reference signal to the analytical instrument.

[0012] U.S. Pat. No. 4,454,752 disclosed a test circuit for use in aphotometric coagulation instrument for plasma samples that verified theelectronic circuitry of the instrument, wherein the rapid rise in clotdensity of a plasma sample may be simulated by a applying to the clotdetection circuitry of the instrument a synthetic waveform thatsimulates the signal that results during clot formation in a reagentplasma mixture. However, this patent did not disclose methods by whichthe proper functioning of an instrument capable of measuring whole bloodcan be analyzed. The disclosed methods are capable of verifying onlythat the clot detection circuits of an photometric plasma coagulationinstrument are performing properly. The patent did not disclose methodsby which other instrument functions such as temperature control, absenceof optical system light leaks, proper detection of sample insertion,etc., may also be verified.

[0013] Verification methods suitable for partially verifying thefunction of certain whole blood coagulation analyzers and unitizedreagent cartridges are also known in the art. For example, U.S. Pat.Nos. 4,948,961 and 5,204,525 disclosed a quality control device for aninstrument with an analysis cartridge constructed so that theinstrument's light passes through the cartridge's internal chamber. Suchsystems have been used for a number of whole blood clinical tests,including whole blood prothrombin time assays when the internal chamberis filled with thromboplastin, and the cessation of red cell movement istracked by light scattering techniques.

[0014] U.S. Pat. No. 5,204,525 disclosed a control device using a liquidcrystal cell interposed between the light source and detector in ananalytical instrument, and a polarizing filter, so that the passage orblock passage of light between the analytical device's light source andlight detector when the voltage to the liquid crystal is modulated.However, neither U.S. Pat. No. 5,204,525 nor U.S. Pat. No. 4,948,961disclosed means by which the temperature control of an analytical devicemay be verified. Although these publications disclosed devices usefulfor monitoring the function of optically transmissive reaction chambersin which the light source passes through the chamber, and which thereaction in question does not alter the wavelength of the light emittedby the instrument's light source, they did not disclose devices usefulfor monitoring the function of fluorescent test strip articles such asthose disclosed in U.S. Pat. No. 5,418,143. In such systems, light ofone wavelength enters a test strip, and excites a fluorescent compoundwhich then emits light that exits the test strip at the same side as thelight source (rather than passing through a reaction chamber), and at adifferent wavelength.

[0015] Another type of control device is found in the BoehringerMannheim “Coaguchek” whole blood prothrombin time analysis devicedisclosed by U.S. Pat. No. 4,849,340. This device uses a disposablereagent cartridge consisting of a chamber with thromboplastin andmagnetic particles. The disposable reagent cartridge is placed in astage in the analysis device, and a blood sample is added. The analysisdevice subjects the reagent cartridge to a varying magnetic field, anddetects the motion of the magnetic particles by the optical interactionbetween the motion of the magnetic particles and a beam of light. Innormal operation, when blood is applied to the disposable reagentcartridge, the magnetic particles are free to move in suspension, andthus provide a high degree of modulation to the optical signal inresponse to the varying magnetic field. As the blood clots in responseto the thromboplastin reagent, the magnetic particles become less ableto move, and thus provide a progressively smaller amount of modulationto the optical signal as time progresses.

[0016] An “electronic control” is provided for the Coaguchek.(Boehringer Mannheim electronic control user manual, 1996) This“electronic control” consists of a separate device consisting of adisposable reagent sized probe that fits in to the reagent stage on theCoaguchek device. The probe contains a magnetic coil pickup, a lightemitting diode, and means to vary the intensity of the response of thelight emitting diode to current generated by the magnetic coil pickup.By using this “electronic control” device, the operator can verify thatthe varying magnetic field generator on the Coaguchek is operatingproperly, and that the optical sensor on the Coaguchek is also operatingproperly. The temperature of the reaction stage, and the performance ofthe optical light source on the Coaguchek, are not tested by thisdevice, however.

[0017] In addition to passive (time unvarying reference signal) andactive (time varying reference signal) verification devices, a thirdtype of verification methodology has been disclosed which incorporatescertain verification systems on to the disposable reagent itself. Thisis disclosed by in U.S. Pat. Nos. 5,591,403 and 5,504,011. U.S. Pat. No.5,591,403 disclosed a reaction chamber cuvette, useful for prothrombintime testing, with multiple conduits. One or more conduits contain thereaction chemistry for the prothrombin time reaction itself, and otherconduits contain control agents useful for assessing certain functionsof the analytical instrument that reads the test cartridge, and the testcartridge itself. Typically one “control” conduit will contain a vitaminK dependent clotting factor concentrate, and a different “control”conduit will contain an anticoagulant. In a properly functioninginstrument, the control conduit with the vitamin K dependent clottingfactor concentrate will initiate a coagulation signal early, and thecontrol conduit with the anticoagulant will initiate a coagulationsignal late. This tests the proper function of those meter detectorelements that read the status of the control conduits. Because thecontrol elements are incorporated into normal prothrombin time reactioncuvette, an independent, non-destructive, test of proper meter functionis not possible with this system.

[0018] Thus, a need exists for an improved verification device. Thisneed and others are addressed by the instant invention.

SUMMARY OF THE INVENTION

[0019] One aspect of the invention is a method for verifying the outputof a system having a radiation source and a radiation detector, saidmethod comprising positioning a reference surface to receive radiationfrom the radiation source and return radiation to the detector; andmodulating at least one of the radiation from the source and theradiation to the detector over time to emulate reflective or radiationcharacteristics of a chemical or biological reaction on the referencesurface.

[0020] A further aspect of the invention is a method for verifying theoutput of a system having a radiation source and a radiation detector,said method comprising positioning a reference surface to receiveradiation from the radiation source and return radiation to thedetector; and modulating at least one of the radiation from the sourceand the radiation to the detector over time in response to temperaturechanges. In some embodiments, the temperature changes are determinedwithin the system. In further embodiments, the temperature changes aredetermined external to the system.

[0021] A further aspect of the invention is an apparatus for use incombination with an analyzer having a radiation source and a radiationdetector, said apparatus comprising a reference surface which producesreturn radiation in response to receiving radiation from the source, andmeans disposed adjacent the radiation surface for modulating at leastone of radiation to the reference surface or radiation from thereference surface. In some embodiments the modulating means modulatesthe radiation over time to emulate reflective or radiationcharacteristics of a chemical or biological reaction on the referencesurface. In further embodiments the modulation means modulates theradiation in response to changes in temperature.

[0022] A further aspect of the invention is an electronically controlledoptical reference device useful for the verification of a clinicalanalytical system having an optical detection apparatus, the referencedevice comprising, an opaque optical reference; an optical shutter;means for controlling the percent exposure of the optical reference tothe optical detection apparatus; optionally, a means to monitor thetemperature of a reaction stage of the clinical analytical system; andan algorithm or method that controls the rate at which the opticalreference is selectively revealed to the optical detection apparatus,said algorithm or method being selected as to simulate the reactionrates of one or more levels of clinical analytes reacting with a testreagent.

[0023] A further aspect of the invention is a verification device usefulfor determining the proper function of an optical, temperaturecontrolled analytical instrument, the device comprising an electronicoptical shutter with an optically active backing, interposed between anoptical signal emitted by the analytical instrument and an opticaldetector mounted on the analytical instrument; a temperature sensor, thesensor contacting a reaction stage on the analytical instrument; andverification device electrodes, the verification device electrodesmaking contact with electrodes on the reaction stage of the analyticalinstrument; wherein the action of the device is initiated by aresistance drop across the verification device electrodes, and whereinthe optical transmission of the liquid crystal shutter is modulated as afunction of time and of the temperature of the reaction stage, wherein arange of levels of enzymatic activity measured by the analyticalinstrument at various operating temperatures is simulated.

[0024] A further aspect of the invention is a verification device usefulfor determining the proper function of an optical, temperaturecontrolled analytical instrument, the device comprising an opticalshutter-fluorescent backing assembly comprising an optical shutterhaving a fluorescent backing placed on one side of the optical shutter;the assembly being interposed between an optical signal emitted by theanalytical instrument and an optical detector mounted on the analyticalinstrument; a thermocouple in contact with a reaction stage on theanalytical instrument; and verification device electrodes, theverification device electrodes making contact with electrodes on thereaction stage of the analytical instrument; wherein the action of thedevice is initiated by a resistance drop across the device electrode,and wherein the fluorescence of the optical shutter-fluorescent backingassembly is modulated as a function of time and of the temperature ofthe reaction stage, wherein a range of levels of enzymatic activitymeasured by the analytical instrument at various operating temperaturesis simulated.

[0025] A further aspect of the invention is an electronically controlledoptical reference device, useful for the verification of an analyticalinstrument having an optical detection apparatus and using opticallyread reagent test strips, the device comprising an opaque opticalreference, which simulates the optical characteristics of a reagent teststrip after reaction with its intended clinical sample; an opticalshutter; a means for controlling the percent exposure of the opticalreference to the optical detection apparatus; and an algorithm or methodthat controls the rate at which the check strip is selectively revealedto the optical detection apparatus, said algorithm or method beingselected as to mimic the reaction rates of one or more levels ofclinical analytes reacting with a reagent test strip.

[0026] A further aspect of the invention is a method for verifying thecorrect performance of a clinical analytical system comprising anoptical detection apparatus, the method comprising contacting theclinical analytical system with an electronically controlled opticalreference device useful for the verification of clinical devices usingoptically read reagent test strips, the reference device comprising, anopaque optical reference, which simulates the optical characteristics ofa reagent test strip after reaction with its intended clinical sample;an optical shutter; means for controlling the percent exposure of theoptical reference to the optical detection apparatus; optionally, ameans to monitor the temperature of the clinical analytical system; andan algorithm or method that controls the rate at which the opticalreference is selectively revealed to the optical device, said algorithmor method being selected so as to mimic the reaction rates of one ormore levels of clinical analytes reacting with a reagent test strip; andanalyzing the optical reference; wherein an expected result of analysisof the optical reference by the clinical analytical system is predictiveof the correct performance of the clinical analytical system.

[0027] A further aspect of the invention is a method for verifying thetemperature control of a clinical analytical system comprising anoptical detection apparatus, the method comprising contacting theclinical analytical system with a verification device useful fordetermining the proper function of an optical, temperature controlledanalytical instrument, the device comprising an electronic opticalshutter with an optically active backing, interposed between an opticalsignal emitted by the analytical instrument and an optical detectormounted on the enzymatic analytical instrument; a temperature sensor,the sensor contacting a reaction chamber on the analytical instrument;and verification device electrodes, the verification device electrodesmaking contact with electrodes on the reaction chamber of the analyticalinstrument; wherein the action of the device is initiated by aresistance drop across the verification device electrodes, and whereinthe optical transmission of the liquid crystal shutter is modulated as afunction of time and of the temperature of the reaction chamber, whereina range of levels of enzymatic activity measured by the analyticalinstrument at a range of operating temperatures is simulated, andanalyzing the optical reference; wherein an expected result of analysisof the optical reference by the clinical analytical system is predictiveof the correct operating temperature of the reaction chamber of theclinical analytical system.

BRIEF DESCRIPTION OF THE DRAWINGS

[0028]FIG. 1 is a diagram depicting an electronic strip emulatorconstructed according to the principles of this disclosure (outside thedotted box), interacting with an exemplary test device (inside thedotted box).

[0029]FIGS. 2A and 2B depict two types of optical shutters. The shutterin FIG. 2A comprises a single shutter element, which can be graduallyvaried from non-transmissive to transmissive. The optical shutter inFIG. 2B comprises numerous shutter “pixel” elements.

[0030]FIG. 3 is a graph depicting the output from the electronicverification device when the analytical device (meter) is at a normaltemperature (37° C.), and at an aberrant temperature (33° C.). Here,Level I mimics a prothrombin time reaction curve obtained from a testsample with a normal prothrombin time value, and Level II mimics theprothrombin time reaction curve obtained from a test sample with anelevated prothrombin time value.

[0031]FIG. 4 depicts an example of a temperature correction algorithmaltering the kinetics of fluorescence development in response to thetime averaged temperature readings from the electronic verificationdevice. The temperature algorithm is selected to match the temperatureresponse of a real reagent test strip.

DESCRIPTION OF THE PREFERRED EMBODIMENT

[0032] The present invention provides a method for verifying the outputof a system having a radiation source and a radiation detector, saidmethod comprising positioning a reference surface to receive radiationfrom the radiation source and return radiation to the detector; andmodulating at least one of the radiation from the source and theradiation to the detector over time to emulate reflective or radiationcharacteristics of a chemical or biological reaction on the referencesurface. In some aspects of the invention, the modulator may operatewithout the reference surface by selectively reflecting light.

[0033] The present invention also provides a method for verifying theoutput of a system having a radiation source and a radiation detector,said method comprising positioning a reference surface to receiveradiation from the radiation source and return radiation to thedetector; and modulating at least one of the radiation from the sourceand the radiation to the detector over time in response to temperaturechanges. In some embodiments, the temperature changes are determinedwithin the system. In further embodiments, the temperature changes aredetermined external to the system.

[0034] The present invention also provides an apparatus for use incombination with an analyzer having a radiation source and a radiationdetector, said apparatus comprising a reference surface which producesreturn radiation in response to receiving radiation from the source, andmeans disposed adjacent the radiation surface for modulating at leastone of radiation to the reference surface or radiation from thereference surface. In some embodiments the modulating means modulatesthe radiation over time to emulate reflective or radiationcharacteristics of a chemical or biological reaction on the referencesurface. In further embodiments the modulation means modulates theradiation in response to changes in temperature.

[0035] The present invention also provides a verification device for aclinical analytical system or instrument. Such a device is also referredto herein as a test strip emulator, a control test simulator, and anelectronically controlled optical reference device. The verificationdevice is typically an electronically controlled optical referencedevice useful for the verification of a clinical analytical systemhaving an optical detection apparatus. The reference device comprised anopaque optical reference or “target”, an optical shutter, and means forcontrolling the percent exposure of the optical reference to the opticaldetection apparatus. The opaque optical reference preferably simulates acalorimetric, fluorescent, or luminescent reagent test strip.Optionally, a means to monitor the temperature of a reaction stage ofthe clinical analytical system is included as part of the device. Thedevice is preferably programed with an algorithm or comprises a methodthat controls the rate at which the optical reference is selectivelyrevealed to the optical detection apparatus. Typically the selectiverevealing of the optical reference is done by exposing the opticalreference to the optical detection apparatus over a specified timeinterval. The algorithm or method is selected so as to simulate thereaction rates of one or more levels of clinical analytes reacting witha test reagent. The algorithm or method can be modified to account forthe temperature of a reaction stage or chamber in the analytical system.

[0036] In some embodiments of the invention, the device of comprises oneor more first electrodes, wherein electrodes contact one or more secondelectrodes on a reaction stage of the clinical analytical system. Theelectrical resistance across the electrodes in the reference device ismodulated to simulate the addition or removal of a disposable reagenttest strip or cartridge or a liquid sample to the clinical analyticalsystem.

[0037] The optical shutter of the device may be electronically operated.Exemplary shutters include but are not limited to a liquid crystalshutter, a magneto-optical shutter, a Faraday effect optical shutter, athermochromic optical shutter, an electrochromic optical shutter, or amicro-mechanical optical shutter. The optical shutter may be dividedinto a plurality of independently or semi-independently controlled pixelelements, such that the optical shutter modulates the intensity of anoptical signal by varying the optical state of the shutter pixels in atime dependent manner.

[0038] In some embodiments, the optical shutter comprises a fluorescentbacking on one side of the optical shutter, and a first optical signalof a first wavelength passes through the optical shutter and interactswith the fluorescence backing, and a fluorescence signal of a secondwavelength passes back through the optical shutter. In furtherembodiments, the optical shutter comprises a colored backing on one sideof the optical shutter, and a first optical signal consisting of a firstspectrum of wavelengths passes through the optical shutter and interactswith the colored backing, and a second optical signal consisting of asubset of the first spectrum of wavelengths passes back through theoptical shutter. The optical shutter may also comprise a luminescentbacking on one side of the optical shutter, with the optical signalcomprising a time increasing or time decreasing luminescence signal.

[0039] In a further feature of the invention, a thermocouple monitorsthe temperature of the reaction stage of the analytical device. Thetransparency of the optical shutter may be modulated as a function oftime and of a thermocouple monitored temperature of the reaction stage,wherein a range of levels of enzymatic activity measured by theanalytical system at various operating temperatures is simulated.

[0040] In some embodiments of the invention, the verification deviceprovides a means to monitor a reagent present and/or blood presentsensor on the clinical analytical device, wherein a stimulus to thesesensors is provided to signal readiness of the meter for testing aclinical sample.

[0041] The instant invention also provides a verification device usefulfor determining the proper function of an optical, temperaturecontrolled analytical instrument. Typically such a device will comprisean electronic optical shutter with an optically active backing,interposed between an optical signal emitted by the analyticalinstrument and an optical detector mounted on the analytical instrument;a temperature sensor, the sensor contacting a reaction stage on theanalytical instrument; and verification device electrodes, theverification device electrodes making contact with electrodes on thereaction stage of the analytical instrument. The action of the device ispreferably initiated by a resistance drop across the verification deviceelectrodes. The optical transmission of the liquid crystal shutter ismodulated as a function of time and of the temperature of the reactionstage, wherein a range of levels of enzymatic activity measured by theanalytical instrument at various operating temperatures is simulated.The reaction stage of the analytical device may be heated.

[0042] In some embodiments the device comprises an opticalshutter-fluorescent backing assembly comprising an optical shutterhaving a fluorescent backing placed on one side of the optical shutter;the assembly being interposed between an optical signal emitted by theanalytical instrument and an optical detector mounted on the analyticalinstrument; a thermocouple in contact with a reaction stage on the isanalytical instrument; and verification device electrodes, theverification device electrodes making contact with electrodes on thereaction stage of the analytical instrument. The action of the device isinitiated by a resistance drop across the device electrode. Thefluorescence of the optical shutter-fluorescent backing assembly ismodulated as a function of time and of the temperature of the reactionstage, wherein a range of levels of enzymatic activity measured by theanalytical instrument at various operating temperatures is simulated.The reaction stage of the analytical device may be heated.

[0043] The instant invention also provides an electronically controlledoptical reference device, useful for the verification of an analyticalinstrument having an optical detection apparatus and using opticallyread reagent test strips. The device comprises an opaque opticalreference, which simulates the optical characteristics of a reagent teststrip after reaction with its intended clinical sample; an opticalshutter; a means for controlling the percent exposure of the opticalreference to the optical detection apparatus; and an algorithm or methodthat controls the rate at which the check strip is selectively revealedto the optical detection apparatus, said algorithm or method beingselected as to mimic the reaction rates of one or more levels ofclinical analytes reacting with a reagent test strip. In a preferredembodiment, the reagent is thromboplastin.

[0044] The instant invention also provides a method for verifying thecorrect performance of a clinical analytical system comprising anoptical detection apparatus using teh devices of the instant invention.In a preferred embodiment, the clinical analytical system is contactedwith an electronically controlled optical reference device useful forthe verification of clinical devices using optically read reagent teststrips. The reference device, for example, comprises an opaque opticalreference, which simulates the optical characteristics of a reagent teststrip after reaction with its intended clinical sample; an opticalshutter; means for controlling the percent exposure of the opticalreference is to the optical detection apparatus; optionally, a means tomonitor the temperature of the clinical analytical system; and analgorithm or method that controls the rate at which the opticalreference is selectively revealed to the optical device, said algorithmor method being selected as to mimic the reaction rates of one or morelevels of clinical analytes reacting with a reagent test strip. Anexpected result of analysis of the optical reference by the clinicalanalytical system is predictive of the correct performance of theclinical analytical system.

[0045] The instant invention also provides a method for verifying thetemperature control of a clinical analytical system comprising anoptical detection apparatus using the devices of the instant invention.In a preferred embodiment, the method comprises contacting the clinicalanalytical system with a verification device useful for determining theproper function of an optical, temperature controlled analyticalinstrument. For example, the device comprises an electronic opticalshutter with an optically active backing, interposed between an opticalsignal emitted by the analytical instrument and an optical detectormounted on the enzymatic analytical instrument; a temperature sensor,the sensor contacting a reaction chamber on the analytical instrument;and verification device electrodes, the verification device electrodesmaking contact with electrodes on the reaction chamber of the analyticalinstrument. The action of the device is initiated by a resistance dropacross the verification device electrodes, and wherein the opticaltransmission of the liquid crystal shutter is modulated as a function oftime and of the temperature of the reaction chamber, wherein a range oflevels of enzymatic activity measured by the analytical instrument at arange of operating temperatures is simulated. An expected result ofanalysis of the optical reference by the clinical analytical system ispredictive of the correct operating temperature of the reaction chamberof the clinical analytical system.

[0046] The verification device of the invention can be provided as aprobe, suitable for insertion into the reaction chamber of acalorimetric, fluorescent, or chemiluminescent test analyticalinstrument, with an opaque colorimetric, fluorescent or luminescenttarget. The reflectance, fluorescence or luminescence of the target ismodulated by an optical shutter. Typically the probe will additionallycontain a temperature sensor, a clock, and means to modulate the opticalexposure of the probes target area according to a preset algorithm. Thepreset algorithm is designed to mimic the response of a normal reagentwith one or more levels of test analyte. The probe may optionallycontain other elements designed to interact with and test other meterfunctional elements, such as a meters “reagent present” and “samplepresent” sensors.

[0047] The optical characteristics of the opaque optical target can varydepending upon the analytical device in question. In one embodiment, thetarget is optically reflective and/or optically colored, so as toeffectively change the distribution of various wavelengths or lightintensity of the target as a function of the state of the opticalshutter. In a further embodiment, the target can made of a fluorescentmaterial, so that the fluorescent intensity of the light detected by theanalytical system's detector varies as a function of the state of theoptical shutter. In a third embodiment, the backing can be luminescent(for example, an electronic luminescent panel), so that the luminescenceseen by the analytical system's luminescence detector varies as afunction of the state of the optical shutter. Although for brevity, thisdiscussion will focus on fluorescent targets, it should be understoodthat the same principles would also apply to calorimetric or luminescentanalytical systems as well.

[0048] A fluorescent target is typically composed of a fluorescentcompound, with absorption and emission characteristics similar to thatof the analytical devices normal fluorescent reagent. The compound willtypically be incorporated into a rigid support matrix. This can be doneby mixing the fluorescent target compound with a suitable supportcarrier, such as acrylic paint, epoxy, or the like. To maximize theoptical signal-to-noise characteristics of the fluorescent target,sufficient quantities of fluorescent compound are added as to completelyinteract with the entire fluorescence optical excitation signal,rendering the target optically opaque. The fluorescent target shifts thewavelength of the excitation signal to a different wavelength, and thefluorescent signal emerges from the side of the target that isilluminated by the excitation wavelength.

[0049] Alternatively, if it is infeasible to make the target opaqueusing large amounts of fluorescent compound, the back of the target maypainted with an opaque backing. The characteristics of this opaquebacking may be selected to maximize the signal-to-noise performance ofthe fluorescent target. If the optical cutoff efficiency of thefluorescent detector's filters to the fluorescence excitationwavelengths is high, the opaque backing could be selected to be of ashiny reflective material. Alternatively, if the optical cutoffefficiency of the fluorescent detector's filters to the fluorescenceexcitation wavelengths is lower, a non-reflective (black) opaque backingmay be chosen to minimize back reflections of the incoming excitationwavelengths to the fluorescence detector.

[0050] In yet another embodiment, the target may be luminescent, andused in a chemiluminescence detecting analytical device that has anoptical detector, but does not contain a light source. The light sourcefor the luminescent target may be provided by variety of conventionalelectrical lighting techniques.

[0051] The optical shutter may be a mechanical or electro-mechanicalshutter, such as an iris as typically used to control exposure intensityin cameras, a series of louvers, or the like. Alternatively, the opticalshutter may be an electro-optical shutter, such as a liquid crystalshutter, a magneto-optical electric shutter, Faraday effect opticalshutter, thermochromic optical shutter, Electrochromic optical shutter,micro-mechanical optical shutter, or other such device. Some exemplaryoptical stutters suitable for the present invention are disclosed, forexample, in U.S. Pat. Nos. 3,649,105; 4,556,289; 4,805,996; 4,818,080;5,050,968; 5,455,083; 5,459,602; and 5,525,430.

[0052] The optical shutter may be composed of a single functionalshutter element, or alternatively it may be composed of many smallerfunctional shutter elements, that collectively act to act to alter theoptical characteristics of the shutter as a whole.

[0053] In an configuration, the shutter is mounted so that lightilluminating the optical (fluorescent) target passes through theshutter. Fluorescent light re-emitted by the optical target may passdirectly to the analytical device fluorescence detector, or optionallypass through the optical shutter on the way to the fluorescencedetector. Alternatively, the optical shutter can be mounted to interactonly with light emitted by the optical target. In still a furtherconfiguration, the optical shutter can interact with light bothilluminating and emitted by the optical target.

[0054] The device may optionally contain means of monitoring thetemperature of the probe near the target area, as well as means ofmodulating the fluorescence signal in response to the temperature of thetarget area. These means may be mechanical, such as a bimetallic stripmechanical temperature sensor device hooked up to a mechanical shutter,chemical, such as a temperature sensitive liquid crystal thermometer, orelectronic, such as a thermistor, thermocouple, or the like. In thepreferred embodiment, the temperature sensor is electronic.

[0055] The means to modulate the target's fluorescence may bemechanical, such as a clockwork mechanism, cam, or the like. The meansmay be controlled by analog electrical circuits, such as simple analogtimers or the like, or the means may be controlled by digital electricalcircuits, such as microprocessors, microcontrollers, and the like. Inthe case of mechanical means, the algorithm encoding the state of thetarget's fluorescence as a function of time is encoded into the designof the mechanical timing elements. In the case of analog electricalcircuits, the algorithm is encoded by properly selected time constants,and the like. In the preferred case of digital microprocessorcontrollers, the algorithm is encoded by a specific program thatcontrols fluorescence as a function of time, and optionally temperatureand other variables.

[0056] To fully validate the analytical system's performance over avariety of sample ranges, the algorithm will ideally simulate thereaction occurring when samples with different relative activity reactwith test reagents. The algorithm may switch from simulating one testlevel to a different test level either in response to user input, orautomatically as the test algorithm runs through a preset series ofvalidation tests.

[0057] The verification device's probe may optionally contain one ormore additional elements that interact with and validate other aspectsof the proper function of the analytical system. For example, the probemay test the function of systems that determine if a cartridge has beenproperly inserted, or systems that determine if sufficient sample hasbeen added. In the case of optical strip insertion or sample additionschemes, the probe may contain additional light emitting or lightblocking devices designed to interact with appropriate optical detectorson the instrument. Alternatively, in the case of electronic stripinsertion or sample addition schemes, such as those disclosed in U.S.Pat. Nos. 5,344,754 and 5,554,531, and in Zweig et. al., BiomedicalInstrumentation & Technology 30: 245-256 (1996), the probe may containone or more electrodes that interact with corresponding electrodesensors on the analytical device, and provide appropriate inputs tosimulate normal activity.

[0058] The verification device may be constructed as a stand-alone,independently powered unit. This may be manually inserted or removed bythe user, or inserted or removed by automated equipment. Alternatively,the verification device may be constructed as an integral part of theanalytical device itself, and may share one or more elements (powersupply, microprocessor time, memory, etc.) with the analytical device.

[0059] A schematic diagram of the verification device of the instantinvention interacting with a meter is depicted in FIG. 1. In thisembodiment, the meter 40 consists of an electrically heated supportstage 27, containing an optical window 30 through which light 22 emittedfrom light source 21 can pass. The meter additionally contains anoptional fluorescence filter 24 and a photodetector 26. In normal use,light 22 travels through the optics window 30 and illuminates afluorescent reagent target. Fluorescent light 23 travels though filter24 and after filtration illuminates photodetector 26. The meter iscontrolled by a microprocessor 29, which initiates test timing inresponse to inputs from strip detect and sample detection electrodes 28.The verification device circuit board additionally contains electrodes14 that interacts with the strip detect and blood detect electrodes 28on the meter's optics block 20. A thermocouple 11 performs anindependent measurement of the temperature of the meter's heated opticsblock 27. The verification device has an optical shutter 12, with abacking 13, and a circuit board 10. A microcontroller 15 is alsoprovided.

[0060] In a preferred embodiment, the verification device has an 8×8pixel liquid crystal optical shutter 12, with an active area of0.375×0.375″, and an exterior size of 0.5″×0.6″, made by Polytronics,Inc. This is placed on a 0.02″ thick circuit board 10, with exteriordimensions of 0.75″, and length of 2″. The exterior circuit board ismade to the same size as a disposable test strip unit normally used inan Avocet Medical prothrombin time detector (see Zweig, et al.,Biomedical Instrumentation & Technology 30, 245-256; FIGS. 3 and 4).

[0061] The optical shutter has a backing 13 consisting of Rhodamine 110mixed with epoxy. The rhodamine 110 retains its normal fluorescenceactivity when mixed with the epoxy, and the epoxy provided a way toaffix the Rhodamine 110 to the back of the optical shutter 12 in durableand permanent manner. The active elements on the circuit board arecontrolled by a Texas Instruments TSS400-S3 sensor signal processor 15,which is a combination microcontroller, liquid is crystal displaydriver, and A/D converter. The TSS400 additionally contains 2 K bytes ofprogrammable EEPROM, which contained the algorithm needed to drive thesystem.

[0062] When turned on (switches not shown), the verification deviceinitially turns all 64 pixels of the 8×8 pixel optical shutter to theopaque mode. Electrodes 14 connecting to the strip present sensors 28 onthe meter's optics block 20 are switched to conducting mode (resistanceis lowered), to allow the Avocet Meter to detect that a test strip isinserted into the optics block. The meter then initiates a warm-upsequence.

[0063] In this preferred embodiment, upon reaching proper temperature,the meter then sends a signal via its sensor electrodes 28 to theverification device electrodes 14 informing the verification device thatthe meter is now warmed up. Alternatively, the meter can signal to theuser that it is ready, and the user can manually transfer thisinformation to the verification device by pressing an electrical switchon the verification device. After the verification device is informedthat the meter is now ready to proceed, the device then reduces theresistance across a second set of electrodes 44, which interact with theblood present sensors 42 on the meter's optics block. Whereas thisresistance drop is normally used to signal the application of blood tothe reagent strip (see, for example, U.S. Pat. No. 5,344,754), in theinstant invention it is used to signal the meter to proceed even thoughno blood has actually been applied.

[0064] In this preferred embodiment, the microcontroller 15 on theverification device consults an algorithm, and selectively switches anincreasingly larger number of pixels on the liquid crystal shutter 12 totransparent mode as a function of a number of variables, including time,the setting of the verification device (e.g. Level I or II control,etc.), and optionally the temperature of the meter's optics stage 27 asmeasured by temperature sensor 11. The meter optical system 20 observesthe fluorescent backing 13 through the optical shutter 12, and observesa progressive increase in overall fluorescence as a function of time.Alternatively, the verification device can progressively alter thevoltage applied to a single element optical shutter element (see FIG.2A) so as to progressively increase the transparency of the singleelement shutter as a function of time and temperature.

[0065] Exemplary algorithms are as follows. The verification devicecontains one or more stored reaction profile algorithms that control thepercent light transmission of the optical shutter as a function of time.A preferred Level I algorithm is:

% fluorescence(% pixels switched on)=100*[0.01*Reactiontime−0.0.35]  Equation 1:

% fluorescence(% pixels switched on)=100*[0.02*Reactiontime−0.9]  Equation 2:

% fluorescence(% pixels switched on)=100*[0.01*Reactiontime−0.05]  Equation 3:

[0066] In a preferred embodiment, every ten seconds (the frequency atwhich the meter took data) for 240 seconds (a typical test duration) theverification device computes all three equations, and chose the resultsbased upon the rule:

[0067] If Equation 1<0, % fluorescence=0;

[0068] If Equation 1>0 and <20%, % fluorescence=Equation 1

[0069] If Equation 1>20% and Equation 2<80%, % fluorescence=Equation 2

[0070] If Equation 2>80% and Equation 3<100%, % fluorescence=Equation 3

[0071] If Equation 3>100%, % fluorescence=100%.

[0072] This produces an “S” shaped reaction profile, shown in FIG. 3,similar to that of a normal prothrombin time (Level I) sample. Incontrast, an exemplary algorithm used for the elevated prothrombin time(Level II) control is:

% fluorescence(% pixels on)=100*[0.007*Reaction time−0.6]  Equation 1,2, and 3:

[0073] The decision tree is the same as the Level I control shownpreviously. This produces a delayed, lower slope, linear curve moretypical of that of a normal Level II sample reaction.

[0074] A typical temperature verification algorithm is as follows. Toverify that the meter's optical stage is being maintained at the propertemperature, the thermocouple on the electronic verification deviceperiodically (every second) performs a temperature measurement. Theresults from each temperature are converted into degrees C. ifnecessary. The degree of deviation of the measured temperature from theideal temperature is used as input into a temperature correctionalgorithm. This temperature correction algorithm advances or retards theschedule of pixel switching on the optical shutter in such a way as tomimic the response of a normal test strip-reagent reaction reacting at adeviant temperature. For a prothrombin time reaction, previous work(Daka et al., Journal of Investigative Surgery 4: 279-290; 1991), hasshown that the optimal reaction temperature for a prothrombin time testis at a reaction time minima, and deviations from this idealtemperature, either positive or negative, prolong the prothrombin timevalue.

[0075] In a normal reagent reaction, the effects of temperature arecumulative. That is, a reaction held at the proper temperature for 95%of the reaction will be only mildly affected if the temperature isslightly deviant during 5% of the reaction. For an exemplary prothrombintime reaction, the effects of non-ideal temperature (either positive ornegative) is to slow down the reaction (the ideal temperature is at areaction time minima). Our own work, as well as the work of Daka et.al., has shown that for the prothrombin time verification devicediscussed by example here, the effects of non-ideal temperatures on theprothrombin time reaction can be approximated by the equation:

Fluorescence(Time,Temp)=Fluorescence(Time−(a*(Temperature Deviation)2),

[0076] where “a” is an experimentally determined coefficient (here takento be 0.1) used to bring the validation device's temperature variationin line with those of an actual reagent test strip.

[0077] The verification device reaction profile, described by the LevelI and II equations above, can be temperature compensated to match anormal reagent reaction profile, reacting at a deviant temperature, bysubtracting a factor proportional to the time weighted temperaturedeviation average from the % fluorescence calculation at each relevanttime point. This delays the onset of fluorescent development. Higherorder polynomial fits, or other temperature compensation functions, canalso be used for these purposes.

[0078] The meter's microcontroller takes a series of fluorescencereadings as for a normal test strip, and interprets the result accordingto its normal test strip algorithm. Previous work (U.S. Pat. No.4,418,141 and the Zweig et. al., supra), has shown that for theprothrombin time example illustrated here, the prothrombin time (PT)time correlates linearly with the time at which the normalizedfluorescence reaction profile first exceeds 10% of its maximum value(Time 10%). Thus by shifting the time at which the verification devicereaction profile first exceeds 10% of its maximum value; the temperaturecompensation algorithm will cause a corresponding shift in theprothrombin time value reported by the meter after reading theverification device.

[0079]FIG. 3 depicts the verification devices' fluorescence profilesreacting according to the Level I and Level II algorithm at a simulatedoptimal temperature of 37° C., and at an aberrant lower temperature.FIG. 4 shows graphically how the verification device's temperaturecorrection algorithm delays the initial onset of the fluorescent signal(Time 10%) for meters operating at aberrant temperatures.

[0080] If the meter is working properly, the answer displayed will bewithin the expected parameters. If the fluorescence detector is workingimproperly, the meter's internal error detection mechanisms will detecta problem (no signal or erratic signal) and display an error code. Ifthe meter's stage is at the incorrect temperature, the deviant patternof pixel switching on the electronic strip emulator causes the meter tooutput an answer outside of the expected parameters. The user can eitherbe instructed to not use the system when this happens, and or the metercan itself examine the results, and automatically lock itself into a“safe” mode to prevent outputting an erroneous answer when a real sampleis used.

EXPERIMENTAL EXAMPLES

[0081] The device constructed according to the preferred embodimentdisclosed above was constructed. A fluorescent backing was provided byfirst mixing 50 mg/ml of Rhodamine-123 in 10 ml of isopropyl alcoholsolution, to produce a 5 mg/ml Rhodamine-123 solution. This was mixed ina 1:3 ratio with “Clear Gloss” acrylic finish (lot 404100, Floquil-PolltS Color Corp., Amsterdam N.Y.) The Rhodamine dye mixed evenly with thisacrylic paint. 10 microliters of this acrylic paint-dye mix was thenapplied to the back of a Polytronics liquid crystal shutter, and allowedto air dry. When dry, the paint formed a clear durable finishencapsulating the Rhodamine-dye. After drying, the back of the liquidcrystal shutter was further coated with black epoxy, forming a moredurable, and light opaque fluorescence backing.

[0082] The verification device was then programmed as described abovefor the preferred embodiment, and tested in a prototype Avocet PT-1000prothrombin time instrument. The instrument responded to theverification device as it would to a normal test strip reacting withsample, and produced appropriate Level I (normal prothrombin time) andappropriate Level II (prolonged prothrombin time) answers.

[0083] All references (including appendices, books, articles, papers,patents, and patent applications) cited herein are hereby expresslyincorporated by reference in their entirety for all purposes.

[0084] While the invention has been described in connection withspecific embodiments thereof, it will be understood that it is capableof further modification, and this application is intended to cover anyvariations, uses, or adaptations of the invention following, in general,the principles of the invention and including such departures from thepresent disclosure as come within known or customary practice in the artto which the invention pertains and as may be applied to the essentialfeatures hereinbefore set forth, and as fall within the scope of theinvention and the limits of the appended claims.

What is claimed is:
 1. An electronically controlled optical referencedevice useful for the verification of a clinical analytical systemhaving an optical detection apparatus, the reference device comprising:an opaque optical reference; an optical shutter; means for controllingthe percent exposure of the optical reference to the optical detectionapparatus; optionally, a means to monitor the temperature of a reactionstage of the clinical analytical system; and an algorithm or method thatcontrols the rate at which the optical reference is selectively revealedto the optical detection apparatus, said algorithm or method beingselected as to simulate the reaction rates of one or more levels ofclinical analytes reacting with a test reagent.